The Agrobacterium VirE2 effector interacts with multiple members of the Arabidopsis VIP1 protein family

T-DNA transfer from Agrobacterium to its host plant genome relies on multiple interactions between plant proteins and bacterial effectors. One such plant protein is the Arabidopsis VirE2 interacting protein (AtVIP1), a transcription factor that binds Agrobacterium tumefaciens C58 VirE2, potentially acting as an adaptor between VirE2 and several other host factors. It remains unknown, however, whether the same VirE2 protein has evolved to interact with multiple VIP1 homologues in the same host, and whether VirE2 homologues encoded by different bacterial strains/species recognize AtVIP1 or its homologues. Here, we addressed these questions by systematic analysis (using the yeast two-hybrid and co-immunoprecipitation approaches) of interactions between VirE2 proteins encoded by four major representatives of known bacterial species/strains with functional T-DNA transfer machineries and eight VIP1 homologues from Arabidopsis and tobacco. We also analysed the determinants of the VirE2 sequence involved in these interactions. These experiments showed that the VirE2 interaction is degenerate: the same VirE2 protein has evolved to interact with multiple VIP1 homologues in the same host, and different and mutually independent VirE2 domains are involved in interactions with different VIP1 homologues. Furthermore, the VIP1 functionality related to the interaction with VirE2 is independent of its function as a transcriptional regulator. These observations suggest that the ability of VirE2 to interact with VIP1 homologues is deeply ingrained into the process of Agrobacterium infection. Indeed, mutations that abolished VirE2 interaction with AtVIP1 produced no statistically significant effects on interactions with VIP1 homologues or on the efficiency of genetic transformation.